TxBR Enabled Reconstruction of Supramolecular Complexes in Intact Microbial Cells
We utilize the Transform-based Tracking, Bundle adjustment, and Reconstruction (TxBR) package to refine our ability to localize specific metabolites, proteins, and functional macromolecular complexes in intact Caulobacter microbial cells imaged with electron microscopic tomography to study cell cycle regulation and cell behavior.
We define the dynamic 3D organization of the chromosome and cell structure within the non-replicating bacterial cell and during the cell cycle while the chromosome is being replicated. To do this, we process data acquired through high-resolution (better than 5 nm3) 3D imaging by EM tomography to determine the configuration of labeled chromosomal loci in the cell and determine the overall spatial distribution of the nucleoid in the cell under different growth conditions. TxBR developed by NBCR and NCMIR, allow for the rapid computation of specialized data generated through the complete projection sampling of bacterial samples (360-degree, total rotation data sets). TxBR includes a specialized alignment system utilizing bundle adjustment to simultaneously estimate the 3D positions of tracked fiducial points within the sample and subsequent transforms of these positions for each projection in the series. Bundle adjustment calculates a complete 3D fiducial model and a 3D-to-2D projection model from the collection of 2D fiducial markers in the EM images. As such, any collection of viewing angles can be used, irrespective of the image acquisition geometry, including the 360-degree tilt geometry utilized for this project. The parallel reconstruction capabilities of this package enhance the throughput of tomographic data collection and allow for extended microbial cell molecular mapping studies.
- Bowman GR, Comolli LR, Gaietta GM, Fero M, Hong SH, Jones Y, Lee JH, Downing KH, Ellisman MH, McAdams HH, Shapiro L. Caulobacter PopZ forms a polar subdomain dictating sequential changes in pole composition and function. Mol Microbiol. 2010;76(1):173-89. doi: 10.1111/j.1365-2958.2010.07088.x.